Presentation on theme: "Specimen Collection and Culture Work-up"— Presentation transcript:
1 Specimen Collection and Culture Work-up Dr Mohammad Rahbar
2 ObjectivesReview collection and transport procedures for wound, stool, respiratory, and genitourinary specimens submitted for microbiological culture.Summarize appropriate algorithms for culture workup of wound, stool, respiratory, and genitourinary specimens submitted for microbiology culture.Discuss the importance of the clinician-laboratory interface
3 Routine Stool Culture Q-Probe Study of 601 institutions Majority of laboratories (99.3%) included Salmonella and Shigella in the routine stool workup96% routinely included Campylobacter30-60% of laboratories surveyed also included other organisms such as Aeromonas, Plesiomonas, Yersinia, Escherichia coli O157, & VibrioValenstein, P, M. Pfaller, and M. Yungbluth The use and abuse of routine stool microbiology. Arch Pathol Lab Med. 120:
4 Role of Rapid Nonculture Screening Tests Fecal Leukocyte TestFecal Lactoferrin AssayGram stain
5 Fecal Leukocyte TestHines, J and I. Nachamkin Effective use of the clinical microbiology laboratory for diagnosing diarrheal diseases. Clin Infect Dis. 23:1292–1301.
6 Fecal Lactoferrin Assay To detect Salmonella, Shigella and Campylobacter spp.The sensitivity ranged from 83 to 93%, and specificity ranged from 61% to 100% 185% sensitive and 79% specific when compared to culture 2As a marker for inflammatory bowel disease (IBD) and irritable bowel syndrome (IBS) 3Significantly higher lactoferrin levels in patients with active and inactive IBD (85.9% sensitivity and 100% specific)Elevated fecal lactoferrin levels – 100% specific in ruling out IBSChoi et. al To culture or not to culture: fecal lactoferrin screening for inflammatory diarrhea. J. Clin. Microbiol. 34:Silletti, R. P. et. al Role of stool screening tests in diagnosis of inflammatory bacterial enteritis in selection of specimens likely to yeild invasive enteric pathogens. J. Clin. Microbiol. 34:Kane, S. V., et. al Fecal lactoferrin is a sensitive and specific marker in identifying intestinal inflammation. Am. J. Gastroenterol. 98:
7 Gram StainThe only role of Gram stain is in the diagnosis of Campylobacter sp.The sensitivity ranges from 66 to 94% with high specificityNo value for detecting other enteric pathogens.Hines, J and I. Nachamkin Effective use of the clinical microbiology laboratory for diagnosing diarrheal diseases. Clin Infect Dis. 23:1292–1301.
8 Number of Specimens Required to Identify a Pathogen Number of Specimens Collected per patientCumulative Percent of Infected Patients DetectedBacteria/Fungi(n = 3349)Parasites(n = 1159)196.991.9299.097.6399.399.8499.499.9Valenstein, P, M. Pfaller, and M. Yungbluth The use and abuse of routine stool microbiology. Arch Pathol Lab Med. 120:
9 Bacteriology Results on Inpatients by Day of Hospital Stay Hospital Day1234>4Total number of specimens collected792468863369202210009Total number of 1st positives, with or without previous specimens374155351949Percentage of total specimens that were patient's 1st positive specimen4.72.31.00.90.5Number of 1st positive specimens from patients for whom previous negative specimens were collected6Valenstein, P, M. Pfaller, and M. Yungbluth The use and abuse of routine stool microbiology. Arch Pathol Lab Med. 120:
10 CAP CHECKLIST Question MIC.22440 PHASE: I Question MIC.22336 PHASE: I Does the laboratory have guidelines (developed with clinicians) for the number and/or timing of collection of stool specimens submitted for routine bacterial testing?Question MIC PHASE: IDoes the final report for routine bacterial stool cultures list the organisms for which the specimen was cultured (e.g., Salmonella, Shigella, Vibrio, etc.)?
11 Acceptable Specimens Fecal sample Fresh Preserved received within 1 to 2 h of passagePreservedBuffered Glycerol Salinerecommended for Salmonella & Shigella but not for Campylobacter or Vibrio sp., unless enriched with CaCl2Modified Carey-Blairgood overall transport mediainappropriate for C. difficile
12 Acceptable Specimens Rectal swabs Duodenal, colostomy or ileostomy contentsstool transport vialsRectal biopsy samplessterile container with a small amount of sterile water
13 Unacceptable Specimens Unpreserved stool samples > 2h oldDry rectal swabs or biopsy specimensMultiple specimens received on the same daySpecimens received from inpatients after the third hospital day, without prior consultation .
14 Workup Guidelines for Salmonella, Shigella, Aeromonas & Plesiomonas species MediaNon-selectiveBAPAeromonas sp., Plesiomonas shigelloides, Yeasts, Staphylococcus aureus, Pseudomonas aeruginosaDifferential enteric agarMacConkey agar, Eosin Methylene Blue (EMB)differentiates lactose-fermenting from non-lactose fermenting coloniesModerately selectiveHektoen Agar, Xylose Lysine Deoxycholate Agar (XLD), or Salmonella Shigella Agarallows growth of Salmonella & Shigella sp. while suppressing the growth of most members of family EnterobacteriaceaeEnrichment brothGN Broth, Selenite Fincreases chances of detecting low numbers of pathogens
15 Workup Guidelines for Salmonella, Shigella, Aeromonas & Plesiomonas species Enteric Screening ProcedureConventionalTSI or KIA, LIA, and UreaCommercial kitslatex agglutinationFull ID of suggestive screening resultsSerological identification of Salmonella and Shigella sp.
16 Campylobacter sp. Most frequently isolated Selective media Campy-Thio (enrichment broth)Campy-BAPSkirrow mediumCampylobacter-cefoperazone-vancomycin-amphotericin (CVA)Identificationgrowth at 42°C, oxidase and catalase positive, Hippurate positiveNalidixic Acid susceptible, Cephalothin resistantLatex agglutination test
17 Campylobacter sp. Rapid Methods ProSpecT® Campylobacter Microplate Assaydetects Campylobacter specific antigens in stool (fresh or in transport medium)utilizes polyclonal anti- Campylobacter specific antigens capture antibodycan be read visually or spectrophometricallyEvaluated in three studiesSensitivities of 80, 89 and 96%Specificities of 99%Flexiable, easy to usereduces cost, reduce turnaround timeCross-reactivity with C. upsaliensis, C. hyointestinalis, or C. helveticus unknownEndtz, H. P., et. al Evaluation of a New Commercial Immunoassay for Rapid Detection of Campylobacter jejuni in Stool Samples. Eur. J. Clin. Microbiol. Infect. Dis. 19:Hindiyeh, M. et. al Rapid Detection of Campylobacter jejuni in Stool Specimens by an Enzyme Immunoassay and Surveillance for Campylobacter upsaliensis in the Greater Salt Lake City Area. J. Clin. Microbiol. 38:Tolcin, R. et. al Evaluation of the Alexon-Trend ProSpecT Campylobacter Microplate Assay. J. Clin. Microbiol. 38:
18 Genus Vibrio Laboratory Diagnosis MediaTCBS (thiosulfate citrate bile salts sucrose) agarsucrose-fermenting (Yellow colonies)V. cholerae, V. alginolyticus, & V. fluvialisnon-sucrose-fermenting (green colonies)V. parahaemolyticus, V. vulnificus (Lactose Fermentor)Susceptible to 150 g of vibriostatic agent (O/129)
19 Escherichia coli Three paradigms by which diarrhea is produced: enterotoxin productionEnterotoxigenic E. coli (ETEC)Enteroaggregative E. coli (EAEC)invasionEnteroinvasive E. coli (EIEC)intimate adherence with membrane signalingEnteropathogenic E. coli (EPEC)Enterohemorrhagic E. coli (EHEC/ STEC)
20 Enterohemorrhagic E. coli Aka Shiga toxin-producing E. coli (STEC)There are at least 100 serotypes of STECOnly one serotype, namely E. coli O157:H7 can be detected in clinical laboratories.Selective media: sorbitol-MacConkey agarconfirm by latex agglutinationVaried geographic distribution - evaluate prevalence for the need of routine workupAvailability of EIA for detection of STEC
21 Yersinia enterocolitica Detection based on conventional methodsSelective media - CIN agardark red “bull’s eye” with a transparent border
22 Clostridium difficile Testing Acceptable specimenUnformed stool specimen unless ileus due to C. difficile is suspected.Rejection criteriaSpecimens that are not liquid or softSpecimens from infants under 1 year old should be discouragedSpecimen more than 24 hours old.Rectal swab specimens“Test for cure” or testing from asymptomatic individualsGerding, D.N. et. al Clostridium difficile-associated diarrhea and colitis.Infect. Control. Hosp. Epidemol. 16:Johnson, S. and D. N. Gerding Clostridium difficile associated diarrhea: a review. Clin. Infect. Dis. 26:
23 Clostridium difficile Testing Culture – most sensitiveSelective medium - Cycloserine-cefoxitin-fructose agarCharacteristic horse-dung smelltypical yellow-green fluorescence under UV lightLimitationsdoes not distinguish between toxigenic and non-toxigenigic strainsdelayed turn-around timeUse of latex agglutination test that detects glutamte dehydrogenase is discouraged
24 Clostridium difficile Testing Cell Culture Cytotoxicity Assay – most specificdetects Toxin BLimitationsRequires 24 to 48 hoursTediousNon-commercial versions are not standardizedEIAs for toxin A or toxins A and BRapidLess sensitive than cell cyotoxicity assayTests that detect only toxin A may miss isolates that are toxin A-B+
26 Wound Cultures: Controversies Is sampling a wound for culture relevant?When and how should wounds be sampled?How should samples be transported?What analysis should be requested?Gram stain only? Culture only? Susceptibility testing?Quantitative cultures?
27 Wounds:Classification AcuteCaused by external damage to intact skinTypesSurgicalBitesBurnsMinor cutsAbrasionsSevere traumaticChronicPrecipitated by predisposing conditions that lead to compromise of dermal/epidermal tissueTypesImpaired venous drainageImpaired arterial supplyMetabolic diseases eg. diabetesBowler PG, et. al Wound microbiology and associated approaches to wound management. Clin Microbiol Rev 14: 244.
28 Wound Infections:Etiology Surgical woundsAerobes: S. aureus, coagulase negative staphylococci, Enterococcus spp. E. coli, P. aeruginosa, Enterobacter spp.Anaerobes: Bacteroides spp., Peptostreptococcus, Clostridium spp.Acute soft tissue infectionsStaph aureus only organism in 30%30-50% mixed aerobes/anaerobes20-30% other eg. Group A streptococci, Clostridium spp.Bite woundsSpecial pathogens: Pasteurella multocida, Capnocytophaga canimorsus, Bartonella henselae, Eikenella corrodensOther mixed aerobes and anaerobes
29 Wound Infections:Etiology Burn woundsPrimarily aerobic organisms: P. aeruginosa, Staphylococcus aureus, E. coli, Klebsiella spp. Enterococcus spp. and Candida spp.Diabetic foot ulcersAerobes: Staph aureus, Streptococcus spp. P. aeruginosa, Enterococcus spp., entericsAnaerobes: Peptostreptococcus, Bacteroides spp., Prevotella spp.Decubitus ulcersMixed aerobic and anaerobic bacteria
30 Wound Cultures For open wounds Clean the wound margins with surgical soap or 70% ethyl or isopropyl alcoholAspirate from the depth of the wound using a sterile syringe and needleAspirated fluid should be sent to the laboratory in an appropriate transport systemAlternatively, a curette may be used to obtain tissue from base of the woundSwabs are strongly discouraged
31 Wound Cultures For closed wounds Prepare site as described for obtaining blood cultureAspirate as much purulent material as possibleTransport in aerobic/anaerobic transport system
32 Wound Cultures: Gram stain ProsUseful in estimating organism load from tissue biopsiesPresence of microorganisms on smear from swabs correlates with > 106 organisms (burns)Facilitates identification of etiologic agent of wound infection following clean surgeryConsPoor correlation seen between Gram stain and culture results from biopsy of diabetic foot infectionsIn mixed infections, little value although presence of leukocytes indicates infection
33 Wound Specimens: Algorithms Three approachesPMN predominanceQ-ScoreQ system
35 Wound CulturesCulture for aerobic and anaerobic bacteria if appropriately collectedGram stain results suggest adequate collection or presence of inflammationTissues or aspirates vs. swabsPrimary plating media: 5% SBA, Choc agar, MacConkey agar; anaerobic plates and thio if appropriately collectedIdentify anaerobes to Genus level onlyPerform susceptibility testing of predominant organisms only
36 Wound Cultures: Extent of Workup Possible approachesUse Gram stain resultWork up organisms seen on stain onlyList othersWork up any potential pathogens to maximum of three, list others present by morphologyWork up any quantity S. aureus, P. aeruginosa, beta hemolytic streptococci, enterics and gram-negative anaerobes
37 Wound Cultures: Examples Gram stain results: (Acceptable)Many neutrophils, no epithelial cellsMany gram positive cocci in clustersMany gram negative bacilliFew morphotypes resembling skin floraWork up (identify and perform susceptibility testing): Gram positive cocci in clusters and gram neg bacilliCulture report: Many S. aureus, many Klebsiella pneumoniae, light aerobic bacteria resembling skin flora
38 Wound Cultures: Examples Gram stain: many neutrophils, few epithelial cells, Gram positive cocci in clusters, Gram positive cocci in chains,Culture grows: many S. aureus, many Group A streptococci, few enteric bacilliWork up: S. aureus, Group A streptococcus: limited ID and no susceptibility on enteric bacilli; susceptibility testing on Group A strep not required
39 Wound Cultures: Examples Gram stain: Many neutrophils, few epithelial cells, multiple morphotypesCulture grows: more than 3 potential pathogensConsider sourceTissue or aspirate ?Contamination likely ?Type of patientMay need to consult with clinician or Infectious Diseases service
40 Q-Score Squamous Epithelial Cells Neutrophils Q-Score = # of potential pathogens (PP)to work upSquamous Epithelial CellsNo Cells1-9/lpf10-24/lpf>25/lpfScore-1-2-3Neutrophils3+1+21+32
41 Workup of Wound Cultures Q-Score SystemGood quality specimen (Q3)Up to 3 organisms can be considered as potential pathogens and worked up (ID/AST)Lower quality specimen (Q2, Q1)More SECFewer organisms are worked up
42 Workup of Wound Cultures Q-Score SystemIf the Q-score is greater than or equals the PP in cultureWorkup all potential pathogensIf Q-Score is less than the PP in cultureLook at the Gram stainWorkup all PP that are seen on GSMorphologically ID othersIf all PP present on GS then only Morph ID all
43 Workup of Wound Cultures Q/2-3-4 SystemCulture workup is based on the # of PP present2PP – ID/AST3PPLook at the Gram stainWorkup two PP if they are seen on GSIf all 3 present on GS then Morph ID4PPMorph ID only
44 Wound Cultures: Example Gram stain: many neutrophils, few epithelial cells, Gram positive cocci in clusters, Gram positive cocci in chains,Culture grows: many S. aureus, many Group A streptococci, few enteric bacilliQ score = 2 [PMN (+3), few epi (-1)]Q/2-3-4 = 3 PPlook at gram stainWork up: S. aureus, Group A streptococcus, Morph ID and no susceptibility on enteric bacilli
49 “ The culture of lower respiratory specimens may result in more unnecessary microbiologic effort than any other type of specimen.” Raymond C Bartlett
50 Lower Respiratory Tract Infections Epidemiology Pneumonia is the sixth leading cause of death in USIncreasing numbers of patients at riskAging populationIncrease in patients with immunocompromising conditionsOvertreatment has lead to resistanceMultidrug resistant Streptococcus pneumoniaeResistance among hospital acquired pathogens such as Acinetobacter, Pseudomonas aeruginosa and others
51 Cumitech 7B:2003 Lower Respiratory Tract Infections 6 contributing authorsMajor sectionsClinical aspects of diseases of LRTSpecimen collectionSpecimen processingInterpretation of bacterial culturesMost common pathogensMethods for implementing changeGuidelines for frequency of testingPublic health issuesReimbursement codes
52 Categories of Lower Respiratory Tract Infections Acute bronchitisCommunity acquired pneumoniaHospital acquired pneumoniaPneumonia in the immunocompromised host
54 Community Acquired Pneumonia Diagnosis Available Test MethodologiesSputum Gram stain and cultureBlood culturesSerologic studiesAntigen detection testsNucleic acid amplification tests
55 Sputum Gram Stain and Culture ProponentsDemonstration of predominant morphotype on Gram stain guides therapyAccuracy is good when strict criteria are usedCheap, so why not?AntagonistsPoor specimen collectionIntralaboratory variability (Gram stain interpretation)Low sensitivity and specificity
56 Sputum Collection Proper patient instruction Food should not have been ingested for 1-2 h prior to expectorationThe mouth should be rinsed with saline or waterPatient should breathe and cough deeplyPatient should expectorate into a sterile containerTransport container immediately to labPerform Gram stain and plant specimen as soon as possible
57 Sputum Gram Stain Screen for acceptability Examine specimen under low power (x 10 objective)Examine 10 representative fieldsSpecimens that show few squamous epithelial cells (< 10/lpf) and many PMNs (> 25/lpf) are acceptableNotify physician of unacceptable samples
61 Sputum Gram Stain Good quality specimens Quantify number and types of inflammatory cellsNote presence of bronchial epithelial cellsConcentrate on areas with WBCs when looking for organismsDetermine if there is a predominant organism (> 10 per oil immersion field)Semiquantitate and report organism with descriptiveIf no predominant organism is present, report “mixed gram positive and gram negative flora”
63 Utility of the Gram Stain in Diagnosis of Pneumonia Roson, B, et. al Clin Infect Dis 31:Prospective studyNon immunocompromised patients hospitalized with CAP1,000 bed hospital in SpainER physicians instructed on sputum collection for Gram stain and cultureSputum collected under supervision of nurse or residentSamples were processed immediatelyScreened for epithelial cellsScreened for predominant morphotype (> 75% of the organisms seen)Sputum planted to blood agar, chocolate agar and MacConkey agarStrictly defined clinical and diagnostic parameters
64 Utility of the Gram Stain in Diagnosis of Pneumonia Roson, B, et. al Clin Infect Dis 31:869-74Results190/533 (35.6%) patients had no sputum sample submitted (these patients were included in the calculations)133/533 (25%) patients had a poor quality specimen210/533 (39.4%) patients had a good quality specimenOverall sensitivity and specificity for pneumococcal pneumonia: 57% and 97%Overall sensitivity and specificity for H. influenzae pneumonia: 82 % and 99%Gram stain gave presumptive diagnosis in 80% of patients who had a good specimen submitted> 95% of patients in whom a predominant morphotype was seen on Gram stain received monotherapy
65 Gram Stain Reports Be as descriptive as possible Moderate neutrophilsModerate Gram positive diplococci suggestive of Streptococcus pneumoniaeFew bacteria suggestive of oral floraKeep report short—avoid line listing of all morphotypes present
66 Sputum and Endotracheal Suction Culture Evaluation Identify and perform susceptibility testing on 2-3 potential pathogens seen as predominant on Gram stainAlpha strep—rule out S. pneumoniaeYeast—rule out Cryptococcus neoformans onlyS. aureus, Gram negative bacilli< normal flora, quantify and limit ID; no susceptibilityAdd comment that organism not predominant on stainID mould, Mycobacteria or Nocardia spp.Modified from Sharp SE, et. Al Cumitech 7B. ASM Press.
67 IDSA Practice Guidelines Diagnostic Tests for CAP OutpatientsEmpiric therapy with a macrolide, doxycycline, or a fluoroquinoloneHospitalized patients with CAPGram stain and culture of sputum2 pretreatment blood culturesStudies for Mtb, Legionella in select patientsRationaleTo improve patient careAdvance knowledge of epidemiologically important organismsPrevent antibiotic abuseReduce antibiotic expenseBartlett JG Clin Infect Dis 31:
68 ATS Guidelines Diagnostic Tests for CAP Empiric therapy for outpatientsMacrolide or tetracyclineHospitalized patients with CAP2 sets of pre-treatment blood culturesPleural fluid Gram stain/culture when appropriateStudies for Legionella, Mtb, fungi in select patientsSputum Gram stain/culture only if resistant or unusual pathogen is suspectedAvoid extensive testingATS Am J Respir Crit Care Med 163:
69 Hospital Acquired Pneumonia Most frequent nosocomial infection (30-33% of cases) among combined medical surgical intensive care units83% are ventilator associatedEtiologic agents Frequency (%)Gram positive cocciS. aureusS. pneumoniaeAerobic gram-neg bacilli 60Pseudomonas aeruginosaEnterobacter sp.Klebsiella pneumoniaeAcinetobacterLegionellaAnaerobesFungiModified from: Carroll KC J Clin Microbiol 40:
70 Hospital Acquired Pneumonia Diagnosis American College of Chest Physicians: Clinical findings are not sufficient for definitive diagnosisQualitative culture or endotracheal sputum has poor predictive valueBronchoscopy is recommended by many pulmonologistsBronchial brushingsBronchial washesProtected specimen brushingBronchoalveolar lavage specimens (BAL)Transbronchial biopsy
72 Respiratory Specimens Bronchoalveolar Lavage (BAL)Samples large area of the lungPerformed using a bronchoscope100 to 250 ml of saline injectedInjected saline along with secretions is collected by aspirationTransthoracic AspirationInvolves percutaneous introduction of a needle directly into the infiltrate
73 Bronchoalveolar Lavage (BAL) Specimen Acceptability Microscopic examination of Gram-stained smearAcceptable<1% of cells present are squamous epithelial cellsUnacceptable>1% of cells present are squamous epithelial cellsThorpe JE et. al Bronchoalveolar lavage for diagnosing acute bacterialpneumonia. J. Infect. Dis. 155:
74 Processing Bronchoscopy Specimens Bronchoscopy brush protectedAerobic bacterial culture and Gram stainAnaerobic bacterial cultureLimited volumeBronchoscopy brush, unprotectedNo anaerobic cultureBronchial washingsUseful only for pneumonia caused by strict pathogensReasonable requests: Mtb, Fungi, Legionella, PneumocystisBronchoalveolar lavageNo anaerobe cultureAmenable to extensive testing for all opportunistic pathogens
75 Interpretation of Quantitative PSB/BAL Dilution MethodQuantify each morphotype present and express as CFU/mlCalibrated Loop MethodQuantify each morphotype present and express as log10 colony count rangesThresholds for significancePSB > 103 CFU/mlBAL > 104 CFU/mlBaselski and Wunderink Clin Micro Rev 7:547
76 Bronchoscopy Samples Quantitative Methods PSB or BAL Baselski and Wunderink Clin Micro Rev 7: vortex s Final dilutionsPlate 0.1 mlChocolate, blood :10Dilute0.1 ml to 9.9 ml salinePlate 0.1 mlChocolate :1000bloodPlate 0.1 mlChocolate :100,000bloodDilute 0.1 ml to 9.9 ml saline
80 GENITAL TRACT SPECIMENS Patients in high risk situations:Patients known to have gonorrheaMale patients with NGU, PGU, epididymitis, and Reiter's SyndromeFemales with mucopurulent cervicitis, urethral syndrome, endometriosis, and salpingitisNeonates born to infected mothersInfertility investigations
81 GENITAL TRACT SPECIMENS Sexually active asymptomatic females who:Are age 25 years or youngerAre pregnantHave evidence of purulent or mucopurulent cervical dischargeExhibit endocervical bleeding, induced by swabbing on examinationHave had a new sex partner in the preceding 2 monthsUse no contraceptives or a non-barrier method for contraception
82 GENITAL TRACT SPECIMENS For FemalesCervical specimens should be collected after removing excess mucous from the cervical os and surrounding mucosaUse a second swab to collect specimen by rotating the swab for 10 to 30 secs. in the endocervical canalCollect vaginal specimens using a speculum without any lubricant
83 GENITAL TRACT SPECIMENS For malesUrethral specimens are collected by inserting a swab 2 to 4 cm. into the urethra and rotating the swab for 2 to 3 seconds
84 GENITAL TRACT SPECIMENS For HSV lesionsFluid from lesions should be aspirated using a syringeSwab can be used to collect vesicle fluid or cellular material from the base of the lesion before crusting and healing have begun
85 Genital Specimens Specimen source Potential Pathogens Primary plating mediaSpecial considerationsCervixChlamydia; GC; herpesSBA, Choc, TM; viral transport media for herpesNAT testing recommended for GC, CTCul-de-sacAnaerobes, GC, CT, entericsSBA, choc, TM,Mac, ana, thioCollect aspirate;Anaerobe transportEndometriumMixed aerobes /anaerobesSurgical biopsy or sheathed catheterVaginaGroup B strep;Mixed aerobes anaerobes BVSBA; LIM or other special brothCulture for Group B strep only; do not culture for BV
86 LABORATORY DETECTION OF BV Clue Cells: vaginal epithelial cells studded with coccobacilliwet mountpH > 4.5Whiff test + (10-20% KOH)Scored gram stainCulture = NOG. vaginalis isolated in > 92% women with BV and 70% asymptomatic womanProbe: AFFIRM (Becton Dickinson)agrees well with high count G. vaginalis
87 Trichomonas vaginalis Common sexually transmitted diseaseDisease associations and adverse outcomesVaginitisUrethritis—men and womenOutcomesAdverse pregnancy eventsAssociated with increased HIV shedding
88 Trichomonas vaginalis Diagnosis Culture- “gold standard”Diamond’s mediaInPouch TV; BioMed Diagnostics, San Jose CA)Barenfanger J, et. al J Clin Microbiol 40:1387.Wet mount—insensitive (~ 50%)Rapid testsXenoStrip-Tv (GenzymeDiagnostics, Inc. San Antonio, Tex.)more sensitive than wet prepless sensitive than cultureuseful as a POC testPillay A, et. al J Clin Microbiol 42:3853.Kurth A, et. al J Clin Microbiol 42:2940.
89 Lab detection (cont.) GLC—no longer used Detection of sialidases (neuraminidases that remove sialic acid from sialogly-coconjugates)In BV, associated with Prevotella and Bacteroides sp.Colorimetric test BVBlue System (Gryphus Diagnostics—91.7% sensitive; 97.8% specificMyziuk L, et. al BVBlue test for diagnosis of bacterial vaginosis. J Clin Microbiol 41:1925.
92 BV Scored Gram Stain ( from Nugent RP 1991;29:297) TYPENumber seen/OIFNone<11-56-30>30Lacto4321Gard/BactCurved GNR
93 Interpretation of Scored Gram Stain 0-3 = Normal4-6 = Intermediatemay indicate trichomoniasis, GC or CTabnormal gram stain, but not consistent with BV7-10 = Consistent with Bacterial VaginosisSignificance of results unknown in pre-menarchal girls or post-menopausal women
95 Who Should be Screened for BV? Women with vaginal symptomsesp. if failed therapyPregnant women at high risk of preterm birthPregnant women with genital symptomsrule out trichomoniasis as wellWomen with gynecologic surgery
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